DESCRIPTION: Many of the most deadly diseases that plague our society evolve quickly, challenging most of our therapeutic strategies. The pressures of evolution will likely result in a variety of pathways for resistance to evolve. Drug resistance can be caused by a change in the balance of molecular recognition events that selectively weakens inhibitor binding but maintains the biological function of the therapeutic target. To reduce the likelihood of drug resistance, the interdependency of the target's function within the context of the biological system in which it exists must be elucidated. Disrupting the therapeutic target's activity is necessary but not sufficient for avoiding resistance. In this collaborative proposal we hypothesize that key pathways and coupled mechanisms confer drug resistance to therapeutic targets. We seek to define the sequence, structural and dynamic, and temporal evolutionary constraints of the interdependency of drug resistance: 1) to recognize the pathways by which resistance occurs and 2) to devise drug design strategies for developing a drug that is robust against resistance. HIV-1 Protease is the perfect case study! HIV evolves very quickly with on average a point mutation introduced in every third genome replicated. HIV protease inhibitors have the potential of being both very potent and robust to resistance. Protease inhibitors are the only HIV inhibitor class that are transition state analogs and can be evolutionarily constrained within the substrate envelope. Inhibitors that leverage both of these characteristics, such as Darunavir (DRV) and similar analogs, have the potential of being robust, nearly resistance proof inhibitors, to drug- nave HIV infected patients. If HIV achieves resistance to these inhibitors it is only through complex pathways and combinations of mutations. Further elucidating these complex pathways will bring us closer to resistance-proof inhibitors. In this project our team will use and develop cutting-edge technology to follow the pathways of drug resistance selection, to elucidate the molecular basis for their interdependent patterns and incorporate these mechanisms into drug design strategies. Together we will make inroads into tackling drug resistance that would be impossible for any of us to individually achieve